Combinatorial chemistry has recently emerged as an effective method for preparing large numbers of chemical compounds for use, e.g., in the discovery of biologically-active agents such as pharmaceutical drugs. In general, combinatorial chemistry is used to prepare collections of compounds, known as libraries, in which all the members of the library share a common core structural element. Such libraries can be prepared by a variety of methods, including solution-phase synthesis and solid-phase synthesis.
Solid-phase synthesis is usually performed by reaction of compounds which have each been immobilized by a covalent linkage to a solid or insoluble support material. The compounds are attached to the support material, which can be a polymeric resin such as polystyrene or polystyrene copolymer, through a linker, and when the synthesis of compounds is complete, the linker can be cleaved to release the final compound or compounds into solution.
The choice of linker for use in a synthesis depends upon the type of synthetic chemistry to be performed and on the conditions to be employed in the synthesis. In general, a linker is preferably inert to the reaction conditions employed during synthesis of the library, so as to avoid loss of compound by premature cleaving of the compound from the solid support. However, the linker should be selected to permit facile cleavage of the compounds from the solid support when the synthesis has been completed.
Numerous linkers have been proposed for use in solid phase combinatorial synthesis (for reviews, see, e.g., F. Guillier et al. Chem. Rev. (2000) 100(6): 2091-2158). Frequently, such linkers are cleavable under either acidic conditions or basic conditions. However, such linkers are not suitable for the preparation of compounds which are not stable to the acidic or basic conditions required for cleavage.
Other linkers have been developed which can be cleaved under more nearly neutral conditions. However, such linkers may be expensive to prepare and in some cases are not compatible with conditions which may be encountered during synthesis of a combinatorial library.
Accordingly, it would be desirable to provide new linkers which are easily and inexpensively prepared and which are compatible with a variety of reaction conditions.
The present invention relates to compositions and methods useful for performing solid-phase chemical synthesis reactions, including synthesis of combinatorial libraries of compounds. The compositions of the invention include linking moieties that can be cleaved under mild conditions, and are suitable for use with a variety of synthetic reaction conditions.
In one aspect, the invention provides a composition comprising an insoluble support covalently attached to a linker moiety. In this aspect, the linker moiety comprises a group represented by the formula (Formula I): 
in which n is 0, 1 or 2; X is CH2, O, S, or NR, in which R is alkyl (which may be substituted) or aryl (which may be substituted); Y is a leaving group, ORxe2x80x2, NHRxe2x80x2, or SRxe2x80x2, in which Rxe2x80x2 is a positively-charged ion, optionally substituted alkyl or optionally substituted aryl; and R1-R5 are each independently selected from the group consisting of H, optionally substituted alkyl or optionally substituted aryl, nitro, alkoxy, aryloxy, cyano, azido, halogen, optionally substituted thioalkyl and optionally substituted thioaryl, and further wherein at least one of R1-R5 is covalently attached to an insoluble support. In certain preferred embodiments, n is 0. In other preferred embodiments, n is 1 and X is CH2. In another preferred embodiment, Y is xe2x80x94OH. In certain embodiments, the insoluble support is agarose; in other embodiments, the insoluble support is polystyrene (including cross-linked polystyrene-divinylbenzene). In other embodiments, the insoluble support can be solubilized in a solvent. In certain preferred embodiments, R2, R3, and R4 are H. In some preferred embodiments, R1 is covalently attached to the insoluble suppor; in other preferred embodiment, R5 is covalently attached to the insoluble support. In certain preferred embodiments, R5 comprises an aminoalkyl group.
In another aspect, the invention provides a method of preparing a chemical compound on an insoluble support. In this aspect, the method comprises the steps of providing a composition of Formula I (in which n, X, Y, and R1-R5 as are defined above); covalently linking a first reactant to the linker moiety to provide a support-bound first reactant moiety; and reacting the support-bound first reactant moiety with a second reactant, under conditions such that a chemical compound on an insoluble support is prepared. In certain preferred embodiments of this method, n is 0. In other preferred embodiments, n is 1 and X is CH2. In another preferred embodiment, Y is xe2x80x94OH, while in other preferred embodiments, Y is a leaving group. In certain embodiments, the insoluble support is agarose; in other embodiments, the insoluble support is polystyrene (including cross-linked polystyrene-divinylbenzene). In other embodiments, the insoluble support can be solubilized in a solvent. In certain preferred embodiments, R2, R3, and R4 are H. In some preferred embodiments, R1 is covalently attached to the insoluble suppor; in other preferred embodiment, R5 is covalently attached to the insoluble support. In certain preferred embodiments, R1 comprises an aminoalkyl group.
In another aspect, the invention provides a method of preparing a chemical compound. This method includes the steps of providing a composition of Formula I; covalently linking a first reactant to the linker moiety to provide a support-bound first reactant moiety; reacting the support-bound first reactant moiety with a second reactant, under conditions such that a chemical compound on an insoluble support is prepared; and cleaving the chemical compound from the insoluble support. In a preferred embodiment, the step of cleaving comprises contacting the chemical compound on an insoluble support with an electrophilic reagent under substantially neutral conditions. In a more preferred embodiment, the electrophilic reagent is I2.
For convenience, certain terms used in the specification and claims are defined below.
The terms xe2x80x9cinsoluble supportxe2x80x9d or xe2x80x9csolid supportxe2x80x9d, as used herein, refer to a solid or insoluble support, commonly a polymeric support, to which a linker moiety can be covalently bonded by reaction with a functional group of the support. Many suitable supports are known, and include materials such as polystyrene resins, polystyrene/divinylbenzene copolymers, agarose, and other materials known to the skilled artisan. It will be understood that an insoluble support can be soluble under certain conditions and insoluble under other conditions; however, for purposes of this invention, a polymeric support is xe2x80x9cinsolublexe2x80x9d if the support is insoluble or can be made insoluble in a reaction solvent and under conditions used to effect the synthesis of chemical compounds on the support, or cleavage of compounds from the support, as described herein.
A variety of supports are known in the art and can be prepared by known techniques. For example, polymers including the carboxylic acid chloride functionality (e.g., xe2x80x94COCl) are known (see, e.g., P. Hodge and D.C. Sherrington, xe2x80x9cPolymer-supported Reactions in Organic Synthesisxe2x80x9d, Chapter 1, (1980)) and can be prepared by treatment of conventional polymer-supported carboxylic acids (e.g., polyacrylic acids) with, e.g., thionyl chloride, oxalyl chloride, and the like. Polymeric supports including sulfonyl chloride functionalities can be obtained by the reaction of a polymer including sulfonic acid moieties with, e.g., thionyl chloride, or by other known methods, for example, the method described in U.S. Pat. No. 5,118,766. Benzyl halide-containing polymers are well known and include chloromethylated polystyrene (e.g., Merrifield resin). Such reactive supports can be reacted with a linker moiety (e.g., through reaction of an amino group of the linker moiety with a resin containing a carboxylic acid chloride) to form a resin-bound linker of the invention. Supports also include materials such as surfaces (e.g., glass or silicon surfaces), beads (such as glass or metallic beads), particles such as microspheres, carbon whiskers or rods, and the like.
In an alternative embodiment, the linker moiety can be attached to a soluble polymeric support such as a polyether moiety (see, e.g., U.S. Pat. No. 5,877,214 to Kim, and references cited therein). Soluble polyether supports have been used for organic synthesis methods in which reactions occur in the solution phase; the polymeric backbone, together with reactive groups, is dissolved in a solvent in which the polymer is soluble. At the conclusion of a given reaction step, a co-solvent (or non-solvent) is added to the reaction mixture, which causes the polymer to become insoluble and to separate from the liquid phase. The polymer, together with the pendant moieties which have been covalently modified, can then be isolated and washed, if desired, as in conventional solid-phase synthesis. Because such polymeric supports are generally handled (e.g., for purposes of purification) through insolubilization as described above, they will be considered to be xe2x80x9cinsolublexe2x80x9d supports for purposes of this invention.
The term xe2x80x9clinkerxe2x80x9d as used herein, refers to a moiety capable of serving as an attachment point for a chemical compound or moiety (i.e., a desired product) that is prepared by solid-phase synthesis. The linker moiety should be capable of retaining the product to the solid support until cleavage of the product from the support is desired, yet permitting cleavage substantially without destruction of the product. Thus, a linker should preferably be substantially inert to reaction conditions used during the synthesis of the product, while being easily cleaved under conditions that do not destroy the product.
The term xe2x80x9ccleavexe2x80x9d or xe2x80x9ccleavingxe2x80x9d, as used herein, refers to the separation of the product from the solid support. In general, a product is cleaved from a solid support when synthesis of the product is complete and the isolation or separation of the product from the solid support is desired. Conditions suitable for cleavage of products from the linkers of the invention are described in more detail, infra.
The term xe2x80x9calkylxe2x80x9d, as used herein, includes cycloalkyl groups and refers to a straight, branched, or cyclic hydrocarbon radical having from 1 to 12 carbon atoms in the carbon chain (3 to 12 ring carbon atoms for cycloalkyl groups). Preferred alkyl groups are lower alkyl groups having 1 to 6 carbons in the carbon chain (3 to 6 ring carbons atoms for cycloalkyls). Alkyl groups also include groups in which the carbon chain is optionally partially unsaturated, as in alkenes and alkynes. Examples of alkyl groups include methyl, ethyl, butyl, isobutyl, sec-butyl, n-octyl, n-decyl, and the like; propenyl, 3-pentenyl, 2-butynyl, and the like. Alkyl groups can also be substituted at one or more positions on the carbon chain with groups such as halogen, hydroxy, amino (including mono- and disubstituted amino groups such as alkyl amino, dialkylamino, arylamino, and diarylamino), C1 to C6 acyloxy, C1 to C6 acyl, C1 to C6 alkoxy, aryloxy, thiol, C1 to C6 alkylthio, thioaryl, alkylcarbonyl, carboxyl, carboxamido, cyano, nitro, and sulfonyl (including alkylsulfonyl, aminosulfonyl and alkoxysulfonyl).
The term xe2x80x9carylxe2x80x9d, as used herein, refers to monocyclic or bicyclic aromatic groups containing from 6 to 12 carbons in the ring portion, preferably 6-10 carbons in the ring portion, such as phenyl, naphthyl or tetrahydronaphthyl. The term xe2x80x9carylxe2x80x9d also includes heteroaryl groups; heteroaryl groups are groups having 5 to 14 ring atoms and 6, 10 or 14 pi electrons shared in a cyclic array; and containing carbon atoms and at least one (optionally two, three, four or five) oxygen, nitrogen or sulfur heteroatoms in the heteroaryl ring system. Examples of heteroaryl groups are: thienyl, benzo[b]thienyl, naphtho[2,3-b]thienyl, thianthrenyl, furyl, pyranyl, isobenzofuranyl, benzoxazolyl, pyrrolyl, imidazolyl, pyrazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, indolizinyl, isoindolyl, 3H-indolyl, indolyl, indazolyl, purinyl, 4H-quinolizinyl, isoquinolyl, quinolinyl, tetrahydroquinolinyl, phthalazinyl, naphthyridinyl, quinazolinyl, phenanthridinyl, acridinyl, perimidinyl, phenanthrolinyl, phenazinyl, isothiazolyl, phenothiazinyl, isoxazolyl, furazanyl and phenoxazinyl groups and the like. Aryl groups can optionally be substituted on one to five positions on the aryl ring system with halogen, hydroxy, amino (including mono- and disubstituted amino groups such as alkyl amino, dialkylamino, arylamino, and diarylamino), C1 to C6 acyloxy, C1 to C6 acyl, C1 to C6 alkoxy, thiol, C1 to C6 alkylthio, thioaryl, alkylcarbonyl, carboxyl, carboxamido, cyano, nitro, and sulfonyl (including alkylsulfonyl, aminosulfonyl and alkoxysulfonyl).
The term xe2x80x9caralkylxe2x80x9d or xe2x80x9carylalkylxe2x80x9d as employed herein by itself or as part of another group refers to C1-6 alkyl groups having an aryl substituent, such as benzyl, phenylethyl or 2-naphthylmethyl.
The term xe2x80x9ccycloalkylxe2x80x9d, as used herein, refers to cycloalkyl groups containing 3 to 9 carbon atoms, preferably 4 to 7 carbon atoms. Typical examples are cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl and cyclononyl.
The term xe2x80x9chalogenxe2x80x9d or xe2x80x9chaloxe2x80x9d, as used herein, refers to chlorine, bromine, fluorine or iodine with chlorine being preferred.
The term xe2x80x9cleaving groupxe2x80x9d as used herein, is art-recognized and refers to a functionality that upon heterolytic bond cleavage departs with an electron pair. In general, a leaving group will be a functionality that can be readily cleaved from a substrate moiety. One of ordinary skill in the art will be able to select a variety of suitable leaving groups. Examples of leaving groups (or, in some cases, the conjugate acid of a leaving group) include, e.g., carboxylates, halogens, tosylates, mesylates, certain alcohols (including phenols such as pentafluorophenol), N-hydroxysuccinimide (NHS), tetrazoles, triazoles, including 1-hydroxybenzotriazole (HOBT), imidazole, azide, ureas (as the tautomeric form, e.g., from the use of carbodiimide activating agents), cyanide and the like.
In one aspect, the invention provides new linkers and insoluble supports for use in the synthesis of chemical compounds, e.g., for solid-phase synthesis of compounds, including combinatorial synthesis. The synthesized compound(s) are readily released from the linkers of the invention under near-neutral conditions, allowing the preparation of compounds which contain sensitive functional groups that might not survive cleavage conditions which require the use of acids or bases to liberate the desired compounds from the solid support.
In one embodiment, the invention provides a composition comprising an insoluble support covalently attached to a linker moiety, the linker moiety comprising a group represented by the formula (Formula I): 
wherein n is 0, 1 or 2; X is CH2, O, S, or NR, in which R is optionally substituted alkyl or optionally substituted aryl; Y is a leaving group, ORxe2x80x2, NHRxe2x80x2 or SRxe2x80x2, in which Rxe2x80x2 is a positively-charged ion (including a proton, an ammonium ion, a metal ion (such as a sodium, potassium, lithium, or other metal ion)), optionally substituted alkyl or optionally substituted aryl; and R1-R5 are each independently selected from the group consisting of H, optionally substituted alkyl or optionally substituted aryl, nitro, alkoxy, aryloxy, cyano, azido, halogen, optionally substituted thioalkyl and optionally substituted thioaryl, and further wherein at least one of R1-R5 is covalently attached to an insoluble support. In a preferred embodiment, n is 0 or 1. In another preferred embodiment, R1 is covalently attached to a solid support and R2-R5 are H.
As described above, the compositions of the invention comprise a support for synthesis covalently bonded to the linker moiety of FIG. 1; the support (which can be a resin (such as polystyrene), agarose, or other supports, e.g., as described herein) can be attached to the linker moiety through a chemical bond or through any moiety (such as a carbon or heterocycle-containing chain or even a cyclic moiety) capable of allowing the linker moiety to a) react with reagents for chemical synthesis and b) release a compound when chemical synthesis is complete. Examples of such attachment moieties are described herein, and can be, for example, C1 to C6 alkyl chain, which alkyl chain can be interrupted by one or two heteroatoms selected from S, O, N, P and Si. In certain preferred embodiments, the alkyl chain is interrupted by one N or O atom.
Certain preferred embodiments of the supports of the invention are described in more detail in the (non-limiting) Examples provided herein. For example, in an embodiment described in Examples 1 and 2, infra, the invention provides a support material of Formula I in which Y is xe2x80x94OLi (the lithium salt of the carboxylate), n is 0, R1-R4 are all hydrogen, and R5 is a linking moiety of the structure xe2x80x94(CH2)3NH-SUP, in which SUP is a solid support (such as polystyrene resin or agarose beads). In the example of Example 3, infra, n is 0, Y is xe2x80x94OH (carboxylic acid), R2, R4 and R5 are all hydrogen, R3 is methyl, and R1 is a bond to a solid support (in this Example, polystyrene resin). In Example 4, infra, n is 0, Y is xe2x80x94OH (carboxylic acid), R2-R5 are all hydrogen, and R1 is a bond to a solid support (in this Example, polystyrene resin). In Example 5, infra, n is 0, Y is xe2x80x94OH (carboxylic acid), R1-R4 are all hydrogen, and R5 is a bond to a solid support (in this Example, polystyrene resin). In Example 6, infra, n is 0, Y is xe2x80x94OH (carboxylic acid), R2-R4 are all hydrogen, R5 is methyl, and R1 is an N-atom-containing attachment to a solid support (in this Example, polystyrene resin). In Example 7, infra, n is 0, Y is xe2x80x94OH (carboxylic acid), R2-R4 are all hydrogen, R5 is methyl, and R1 is an O-containing attachment to a solid support (in this Example, polystyrene resin). In Example 8, infra, n is 1, X is xe2x80x94CH2xe2x80x94, Y is xe2x80x94OH (carboxylic acid), R2-R5 are all hydrogen, and R1 is a bond to a solid support (in this Example, polystyrene resin).
Compositions of Formula I can be prepared according to a variety of methods, including those described herein and other methods which are known to one of ordinary skill in the art. In general, the compositions can be prepared in at least two ways: (A) by preparing the linker moiety and attaching it to a suitable support; and (B) preparing a linker moiety that can be polymerized to form a linker-support composition of the invention.
In general, a linker moiety of the invention will include a carbonyl group spaced, by a distance of 2, 3 or 4 atoms (more preferably, 2 or 3 atoms), from a carbonxe2x80x94carbon double bond. This arrangement permits the cleavage (or release) of a product from the linker by treatment of the linker with an electrophilic reagent. Without wishing to be bound by any theory, it is believed that an oxygen atom of the carbonyl group can, through intramolecular attack upon the carbonxe2x80x94carbon double bond, form a 5-, 6-, or 7-membered ring (a lactone) with concomitant release of the product from the linker. Certain chemical protecting groups are known which are believed to be cleaved through this mechanism; see, e.g., Madsen, R. et al. J. Org. Chem. (1995) 60:7920; Guo, M J et al. Bioorg. Med. Chem. Lett. (1998) 8:2539-2544; and see generally Greene, T. W. and Wuts, P. xe2x80x9cProtective Groups in Organic Synthesisxe2x80x9d 3rd Ed. (1999), New York: John Wiley and Sons. Conditions suitable for cleavage of products from such linkers are described in more detail, infra.
A. Attachment of a Linker Moiety to a Support
A linker moiety of the invention can be prepared in a variety of ways. For example, in a preferred embodiment, a side-chain of the linker (e.g., R4 or R5 of Formula I) can be provided with a reactive group suitable for covalent attachment to a solid support. For example, as described in Examples 1 and 2 herein, an aminopropyl sidechain can be attached to a solid support by formation of an amide bond, e.g., by reaction of the nitrogen atom of the aminopropyl group with an activated carboxyl group of the solid support.
In another embodiment, a precursor of the linker moiety (e.g., a group which corresponds to R1 of Formula I) can be provided with a reactive moiety such as an aldehyde or ketone group, which can be reacted with a solid-supported Wittig reagent to covalently link the linker to the solid support. For example, ethyl levulinate (which includes a ketone group) can be reacted with a resin-bound phosphonium salt to create a resin-bound linker (see Example 3, infra). Thus, a pre-formed linker moiety (or a precursor thereof) can be covalently secured to a solid support using a single-step reaction, which can, as described herein, serve both to tether the linker moiety to the support and to provide the olefinic bond necessary for linker cleavage by electrophilic activation. In an analogous fashion, a phosphonium salt can be reacted with a resin-bound aldehyde group to form a composition of the invention (see Example 4, infra).
Another method for attachment of the linker moiety to a solid support involves the use of radical reactions to form a covalent bond. For example, a linker moiety which contains a carbonxe2x80x94carbon double bond (such as a terminal alkenyl moiety or a styryl moiety) can be linked to a solid support, such as a resin, which also contains a carbonxe2x80x94carbon double bond, by using a radical initiator (such as 2,2xe2x80x2-azobisisobutyrylnitrile (AIBN)). Such a radical reaction will result in covalent attachment of the linker moiety to the support; the amount of linker attached to the support can be controlled by varying the number of reactive sites on the resin.
Another method for providing a linker covalently attached to a support is to copolymerize a monomeric precursor linker moiety (e.g., a free linker moiety which includes a terminal alkenyl or styryl group) with one or more monomeric blocks such as styrene or divinylbenzene, preferably under conditions known for the production of copolymer resins (such as free-radical polymerization). The copolymerization of the monomer units will result in incorporation of the linker moiety into the resin product, thereby providing the linker covalently linked to the support.
B. Attachment of Compounds to the Linker
Once the linker moiety is prepared and covalently bound to the support (e.g., a resin), the linker/support can be used for synthesis of organic compounds. In general, the synthesis will require the attachment of a reactive moiety to the linker, with subsequent modification of the reactive moiety to produce the desired product(s). Thus, for example, an amine-containing moiety can be coupled to the linker, e.g., through a carboxylate functionality on the linker, by well-known methods for forming an amide bond. For example, in the synthetic scheme shown for Example 9, below, piperazine is reacted with an activated carboxylic acid (as the NHS ester) to provide a piperazine-based scaffold for further synthetic manipulation. Many other compounds can be attached to the linker in a similar manner; for example, a molecule which contains a hydroxyl group can be esterified to the linker carboxylate, through an ester linkage, to provide a scaffold for synthesis of compounds; similarly, a thiol-containing compound can be attached to the linker (e.g., through a thioester group) to provide a template for synthesis. One of ordinary skill in the art will be aware of other methods for attaching compounds to the linker to provide a functionalized material useful for synthesis.
The functionalized support can then be used for solid-phase synthesis, e.g., according to methods well known to the ordinarily-skilled artisan; also see infra.
C. Detachment of Products from the Linker/Support
Once a desired compound (or compounds) has been prepared on the solid support/linker of the invention, the desired compounds can be released from the support, if desired, e.g., to provide compounds in solution for further purification or testing. It will be appreciated, however, that the compounds need not be released from the support, e.g., if it is desired to screen the compounds for a specific activity while the compounds are still attached to the linker/support.
To release the compounds, it is preferable to use relatively mild conditions, to avoid decomposition or undesired functionalization of the compounds. As described above, it is believed (without wishing to be bound by theory) that an oxygen atom of the carbonyl group of the linker portion can, through intramolecular attack upon the carbonxe2x80x94carbon double bond of the linker, form a 5- or 6-membered ring (or, in certain embodiments, a 7-membered ring) with concomitant release of the product from the linker upon hydrolysis. Thus, any reagent or condition that promotes such intramolecular attack can be employed to release compounds from the linker. Examples of reagents suitable for promoting cleavage are: iodine (I2), bromine, iodine monochoride, N-bromosuccinimide, N-iodosuccinimide, mercuric chloride or other mercury(II) compounds, certain protic or Lewis acids, and the like. Such cleavage reactions will typically be performed by suspending the functionalized support, with the attached compound(s), in a suitable solvent with addition of the cleaving reagents. As noted above, a suitable nucleophilic reagent or solvent (such as water) should be added to facilitate the cleavage process.
Preferably, the release of compounds is performed under substantially neutral conditions (e.g., strong acids or bases are not used). In preferred embodiments, release of the compounds is performed at a pH in the range of 6.0-8.0, more preferably 6.5 to 7.5.
Once the compound has been released from the linker and solid support as described above, the desired compounds can be recovered by standard means. For example, when the cleavage reaction is performed on a suspension of resin in a solvent, the desired compounds will be released into, and preferably dissolved or suspended in, that solvent. Once the cleavage process is substantially complete, the resin can be separated from the liquid phase, e.g., by filtration, and the desired compound(s) can be recovered from the liquid phase by well-known techniques such as evaporation, crystallization, extraction, chromatography (including column chromatography, high-performance liquid chromatography (HPLC), and other chromatographic techniques), and other purification and isolation methods which will be apparent to one of ordinary skill in the art.
One advantage of the resin-bound linkers of the invention is that, in certain embodiments, the resin/linker can be regenerated and recycled after a synthesis is complete and the desired product has been released from the resin. Thus, for example, after a product has been released using iodine to promote intramolecular ring formation (i.e., the released resin will include a lactone ring) as described above, the spent resin (containing a vicinal acyloxyiodoalkyl functionality) can be recovered. The spent resin can be recycled for further use by, for example, treatment with allyltrimethylsilane and tin(IV) chloride or titanium(IV) chloride to ring-open and reduce the vicinal acyloxyiodoalkyl functionality (see, e.g., Yachi, K. et al., Tet. Lett. 38(29):5161-5164 (1997)) to provide a carbonxe2x80x94carbon double bond. Alternatively, the lactone ring of the spent resin can be cleaved (e.g., by basic hydrolysis) to yield an iodohydrin (with a free carboxylate group also liberated). The iodohydrin can then be converted back to a double bond by reduction with a reagent such as zinc or magnesium metal. The resulting regenerated resin can then be used in a further synthesis by coupling reagents to the carboxylate group as described above.
In general, the methods of the invention involve the attachment of chemical compounds or moieties to a solid-supported linker group as described above. Such chemical compounds or moieties can then be modified by stepwise reaction under a selected reaction scheme until a desired product is obtained. The desired compound can then be cleaved from the solid support under mild conditions which do not significantly destroy or modify the desired compound.
In one embodiment, the invention provides a method of preparing a chemical compound on an insoluble support. The method comprises the steps of providing a composition of Formula I; covalently linking a first reactant to the linker moiety to provide a support-bound first reactant moiety; and reacting the support-bound first reactant moiety with a second reactant, under conditions such that a chemical compound on an insoluble support is prepared.
In this embodiment, the first reactant is preferably an amine, an alcohol, or a thiol (or a conjugate base of any of these). For example, an amine can be attached to the linker moiety by formation of an amide bond with a carboxylate moiety of the linker (e.g., where Y is xe2x80x94ORxe2x80x2 of Formula I), preferably through the use of an active ester or coupling reagent as is well known in the art. Similarly, an alcohol can be attached through an ester moiety by use of a suitably-functionalized linker (e.g., a linker of Formula I in which Y is a leaving group such as xe2x80x94Cl) or by use of a coupling agent (e.g., where Y of the linker of Formula I is xe2x80x94ORxe2x80x2).
The step of reacting the support-bound first reactant moiety with a second reactant can include the use of a wide variety of synthetic reactions, such as those described herein or known to the ordinarily-skilled artisan.
The chemical compound can be screened for a desired activity by screening the compound on the bead according to methods known in the art (see, e.g., E. M. Gordon et al. J. Med. Chem. (1994) 37:1385-1401, and references cited therein). Alternatively, the chemical compounds, once prepared, can be cleaved from the support (e.g., as described herein) and screened in solution.
In another embodiment, the invention provides a method of preparing a chemical compound, the method including the steps of providing a composition of Formula I; covalently linking a first reactant to the linker moiety to provide a support-bound first reactant moiety; and reacting the support-bound first reactant moiety with a second reactant, under conditions such that a chemical compound on an insoluble support is prepared; and cleaving the chemical compound from the insoluble support.
The step of cleaving the chemical compound from the support can be performed as described previously. In certain embodiments, the step of cleaving comprises contacting the chemical compound on an insoluble support with an electrophilic reagent under substantially neutral conditions; in a preferred embodiment, the electrophilic reagent is I2.
The reactions of the present invention may be performed under a wide range of conditions, though it will be understood that the solvents and temperature ranges recited herein are not limitative and only correspond to a preferred mode of the process of the invention.
A variety of synthetic methods are compatible with the compositions of the invention. For example, synthetic reactions, such as amidation, nucleophilic substitution, cycloadditions, aldol reactions, and the like, can be used to prepare a wide variety of compounds on the solid support (see, e.g., B. A. Lorsbach and M. J. Kurth, Chemical Reviews (1999) 99(6): 1549-1582; R. E. Sammelson and M. J. Kurth, Chemical Reviews (2001) 101(1): 137-202; P. H. Seeberger and W.-C. Haase Chemical Reviews (2000) 100(12): 4349-4394; and R. G. Franzxc3xa9n J. Comb. Chem. (2000) 2(3): 195-214; and references cited therein). Deprotection steps can be performed if necessary; however, as with all synthetic reactions to be performed, it is preferred that deprotection steps are compatible with the linker moiety (i.e., such reaction steps do not destroy the linker, the solid support, or the compounds being synthesized on solid support). For example, a synthetic step which involves the use of I2 could cause premature release of compounds from the solid support, which would likely result in lower yields of the desired product when synthesis is complete.
In general, it is desirable that reactions are run using mild conditions that will not adversely affect the substrate, the nucleophile, the intermediates, or the product. For example, the reaction temperature influences the speed of the reaction, as well as the stability of the reactants and catalyst. The reactions will usually be run at temperatures in the range of xe2x88x9278xc2x0 C. to 100xc2x0 C., more preferably in the range xe2x88x9220xc2x0 C. to 50xc2x0 C. and still more preferably in the range xe2x88x9220xc2x0 C. to 25xc2x0 C.
In general, the reactions according to the invention will be performed using a liquid phase, e.g., the reaction can take place on a support dissolved or suspended in a liquid phase. The reactions may be run in an inert solvent, preferably one in which at least one of the reaction ingredients (such as the support or, more preferably, at least one of the reagents) is substantially soluble. Suitable solvents include ethers such as diethyl ether, 1,2-dimethoxyethane, diglyme, t-butyl methyl ether, tetrahydrofuran (THF) and the like; halogenated solvents such as chloroform, dichloromethane, dichloroethane, chlorobenzene, and the like; aliphatic or aromatic hydrocarbon solvents such as benzene, toluene, hexane, pentane and the like; esters and ketones such as ethyl acetate, acetone, and 2-butanone; polar aprotic solvents such as acetonitrile, dimethylsulfoxide, dimethylformamide (DMF) and the like; or combinations of two or more solvents. In certain embodiments, the use of solvents such as water or alcohols (such as methanol, ethanol, propanol, t-butanol, and the like), either alone or in mixtures with other solvents, may be acceptable.
In certain embodiments it is preferable to perform the reactions under an inert atmosphere of a gas such as nitrogen or argon.
The invention also contemplates the synthesis of libraries or collections of chemical compounds. Combinatorial libraries of compounds can be prepared on solid supports by a variety of methods, some of which are known in the art (see, e.g., F. Guillier et al. Chem. Rev. (2000) 100(6): 2091-2158; E. M. Gordon et al. J. Med. Chem. (1994) 37:1385-1401). In general, preparation of a library involves the use of a plurality of supports (e.g., a plurality of resin beads, plastic xe2x80x9cpinsxe2x80x9d, resin crowns, a plurality of spatially-addressable points on a solid surface, etc.); by varying the reagents used to prepare chemical compounds on each of the plurality of supports, a variety of compounds can be prepared. For example, a plurality of supports (e.g., resin beads), each comprising a composition of Formula I, can be derivatized by linking a first reactant to the linker moiety of each support (e.g., resin bead) to provide a plurality of supports having a support-bound first reactant moiety; and each of the plurality of supports having a support-bound first reactant moiety can then be reacted with a second reactant, under conditions such that a chemical compound on an insoluble support is prepared on each support. The process can be continued, if desired, with third, fourth, etc. reactants to provide the desired chemical compounds. The first, second or subsequent reactants need not be the same for each support; generally, at least one of the reactants will differ between at least two supports, such that at least two different chemical compounds are prepared.
For example, in Example 9, infra, a plurality of supports (6) are reacted with piperidine and p-acetylbenzoic acid to provide a plurality of supports having a support-bound first reactant moiety; the derivatized supports are then divided into five separate portions and reacted with five different aldehydes to provide five different chalcone compounds on solid support. The five portions of support were then recombined and reacted with further reagents (an isatin and an amino acid) to produce five different spirocyclic compounds on the mixed resin supports. The compounds were then cleaved from the resin to provide a mixed library of compounds.
It will be appreciated, however, that in the above-described example, by varying the isatin reagent or the amino acid reagent, a combinatorial library of many members could be provided. For example, use of 5 aldehydes, 5 isatins, and 10 amino acids would provide a library having 250 compounds). Thus, the methods of the invention can be used to prepare libraries having at least 5, 10, 50, 100, 250, 500, 1000, 5000, or 10000 (or even more) compounds.
The following Examples are offered by way of illustration and not limitation.